Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP
This paper presents a universal point-of-care system for fully automated quantification of human T-cell lymphotropic virus type 1 (HTLV-1) proviral load, including genomic RNA, based on digital reverse RNA transcription and c-DNA amplification by MD LAMP (mediator displacement loop-mediated isotherm...
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doaj-4a5b12acd76d4d2a9e78791e90dfe3372021-02-06T00:05:51ZengMDPI AGMicromachines2072-666X2021-02-011215915910.3390/mi12020159Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMPLisa Becherer0Jacob Friedrich Hess1Sieghard Frischmann2Mohammed Bakheit3Hans Nitschko4Silvina Stinco5Friedrich Zitz6Hannes Hofer7Giampiero Porro8Florian Hausladen9Karl Stock10Dominik Drossart11Holger Wurm12Hanna Kuhn13Dominik Huber14Tobias Hutzenlaub15Nils Paust16Mark Keller17Oliver Strohmeier18Simon Wadle19Nadine Borst20Roland Zengerle21Felix von Stetten22Hahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyMast Diagnostica GmbH, Feldstraße 20, 23858 Reinfeld, GermanyMast Diagnostica GmbH, Feldstraße 20, 23858 Reinfeld, GermanyDepartment of Virology, Pettenkoferstraße 9a, Max von Pettenkofer-Institute, 80336 Munich, GermanyDepartment of Virology, Pettenkoferstraße 9a, Max von Pettenkofer-Institute, 80336 Munich, GermanyE.L.T. Kunststofftechnik & Werkzeugbau GmbH, Weidenweg 339, 8240 Friedberg, AustriaE.L.T. Kunststofftechnik & Werkzeugbau GmbH, Weidenweg 339, 8240 Friedberg, AustriaDatamed srl, Via Achille Grandi 4/6, 20068 Peschiera Borromeo MI, ItalyInstitut für Lasertechnologien in der Medizin und Meßtechnik, University of Ulm, Helmholtzstraße 12, 89081 Ulm, GermanyInstitut für Lasertechnologien in der Medizin und Meßtechnik, University of Ulm, Helmholtzstraße 12, 89081 Ulm, GermanyInstitut für Lasertechnologien in der Medizin und Meßtechnik, University of Ulm, Helmholtzstraße 12, 89081 Ulm, GermanyInstitut für Lasertechnologien in der Medizin und Meßtechnik, University of Ulm, Helmholtzstraße 12, 89081 Ulm, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyHahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, GermanyThis paper presents a universal point-of-care system for fully automated quantification of human T-cell lymphotropic virus type 1 (HTLV-1) proviral load, including genomic RNA, based on digital reverse RNA transcription and c-DNA amplification by MD LAMP (mediator displacement loop-mediated isothermal amplification). A disposable microfluidic LabDisk with pre-stored reagents performs automated nucleic acid extraction, reaction setup, emulsification, reverse transcription, digital DNA amplification, and quantitative fluorogenic endpoint detection with universal reporter molecules. Automated nucleic acid extraction from a suspension of HTLV-1-infected CD4+ T-lymphocytes (MT-2 cells) yielded 8 ± 7 viral nucleic acid copies per MT-2 cell, very similar to the manual reference extraction (7 ± 2 nucleic acid copies). Fully automated sample processing from whole blood spiked with MT-2 cells showed a comparable result of 7 ± 3 copies per MT-2 cell after a run time of two hours and 10 min.https://www.mdpi.com/2072-666X/12/2/159human T-cell lymphotropic virus 1digital loop-mediated isothermal amplificationpoint-of-care testingcentrifugal microfluidicsproviral load measurements |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Lisa Becherer Jacob Friedrich Hess Sieghard Frischmann Mohammed Bakheit Hans Nitschko Silvina Stinco Friedrich Zitz Hannes Hofer Giampiero Porro Florian Hausladen Karl Stock Dominik Drossart Holger Wurm Hanna Kuhn Dominik Huber Tobias Hutzenlaub Nils Paust Mark Keller Oliver Strohmeier Simon Wadle Nadine Borst Roland Zengerle Felix von Stetten |
spellingShingle |
Lisa Becherer Jacob Friedrich Hess Sieghard Frischmann Mohammed Bakheit Hans Nitschko Silvina Stinco Friedrich Zitz Hannes Hofer Giampiero Porro Florian Hausladen Karl Stock Dominik Drossart Holger Wurm Hanna Kuhn Dominik Huber Tobias Hutzenlaub Nils Paust Mark Keller Oliver Strohmeier Simon Wadle Nadine Borst Roland Zengerle Felix von Stetten Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP Micromachines human T-cell lymphotropic virus 1 digital loop-mediated isothermal amplification point-of-care testing centrifugal microfluidics proviral load measurements |
author_facet |
Lisa Becherer Jacob Friedrich Hess Sieghard Frischmann Mohammed Bakheit Hans Nitschko Silvina Stinco Friedrich Zitz Hannes Hofer Giampiero Porro Florian Hausladen Karl Stock Dominik Drossart Holger Wurm Hanna Kuhn Dominik Huber Tobias Hutzenlaub Nils Paust Mark Keller Oliver Strohmeier Simon Wadle Nadine Borst Roland Zengerle Felix von Stetten |
author_sort |
Lisa Becherer |
title |
Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP |
title_short |
Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP |
title_full |
Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP |
title_fullStr |
Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP |
title_full_unstemmed |
Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP |
title_sort |
point-of-care system for htlv-1 proviral load quantification by digital mediator displacement lamp |
publisher |
MDPI AG |
series |
Micromachines |
issn |
2072-666X |
publishDate |
2021-02-01 |
description |
This paper presents a universal point-of-care system for fully automated quantification of human T-cell lymphotropic virus type 1 (HTLV-1) proviral load, including genomic RNA, based on digital reverse RNA transcription and c-DNA amplification by MD LAMP (mediator displacement loop-mediated isothermal amplification). A disposable microfluidic LabDisk with pre-stored reagents performs automated nucleic acid extraction, reaction setup, emulsification, reverse transcription, digital DNA amplification, and quantitative fluorogenic endpoint detection with universal reporter molecules. Automated nucleic acid extraction from a suspension of HTLV-1-infected CD4+ T-lymphocytes (MT-2 cells) yielded 8 ± 7 viral nucleic acid copies per MT-2 cell, very similar to the manual reference extraction (7 ± 2 nucleic acid copies). Fully automated sample processing from whole blood spiked with MT-2 cells showed a comparable result of 7 ± 3 copies per MT-2 cell after a run time of two hours and 10 min. |
topic |
human T-cell lymphotropic virus 1 digital loop-mediated isothermal amplification point-of-care testing centrifugal microfluidics proviral load measurements |
url |
https://www.mdpi.com/2072-666X/12/2/159 |
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